**3.2 Aphid toxicity**

Crude vegetable water from olives has been shown to be very toxic to *A. pomi*, their effects are amplified with concentration (**Figure 3**). The response of the treated individuals varies significantly between stages (F = 59.93, df = 4, 141, P < 0.001) and according to the concentration (F = 871.75, df = 6, 141, P < 0.001). Compared to the negative control, crude OMW significantly affect the survival of the different stages of the green apple aphid, the percentages of mortalities are approximately 4 to 38 times higher than the control. Whereas all stages have a comparable mortality rate in untreated lots (P > 0.05). On the other hand, compared to the positive control (Imidacloprid), only the high concentration (80 mg/L) which makes it possible to induce statistically comparable mortalities, the other concentrations cause mortalities markedly lower than this,


**Table 2.**

*Relationship between concentration (x in mg of OMW/L of distilled water) and mortality (y in %) of Aphis pomi treated with crude olive mill wastewater during 48 h.*


*a L = nymph.*

*b Probit model = slope – constante, SE: standard error.*

*c 95% lower and upper confidence limits are shown in parenthesis expressed in mg of OMW/L of distilled water. d Df: degree of freedom.*

*e Probaility.*

#### **Table 3.**

*Toxicity parameters calculated for Aphis pomi exposed during 48 hours to crude OMW.*

*Valorization of Olive Mill Wastewater in the Control of* Aphis pomi *De Geer 1773… DOI: http://dx.doi.org/10.5772/intechopen.100016*

**Figure 4.**

*Toxic activity of crude OMW against the different stages of Aphis pomi after 48 hours of treatment (on the x-axis are presented the concentrations instead of their logarithms).*

P < 0.05 (**Figure 3**). With regard to the same concentration of OMW, the aphid's response varies depending on the stage. With 5 mg /L, the L1 and L2 are affected by the same mortality rate (P > 0.05); the same is true for L3 and L4 or L4 and adults compared in pairs. The same pattern is obtained with 10 mg/L. Treated with 20, 40 or 80 mg/L, the L1 and L2 were shown to be much more vulnerable than the other stages, P < 0.05.

#### **Figure 5.**

*Classification of the different stages according to their sensitivity to OMW (unweighted averages of lethal concentrations).*

Moreover, the relationship linking the mortality of the stages of the aphid to the concentrations of OMW makes it possible to note that for each stage of the green apple aphid, the mortality therefore depends linearly and positively on the concentration of OMW (**Table 2**). In terms of corrected mortality, i.e., due exclusively to crude OMW, the percentages of dead insects vary from around 2 to 98% depending on the concentration and the stage considered with strong variations (Coefficients of variation = 4–70%).

Furthermore, the observed data fit well to the log-probit model and no statistically significant deviation from the regression equation was detected, P > 0.05 (**Table 3** and **Figure 4**). Lethal concentrations vary depending on the stage of the aphid considered; in fact, the extreme LC50 and LC95 vary from approximately 19 to 55 and 65 to 147 mg of OMW/L of distilled water, respectively. With regard to the values of the LC50, the tolerance of *A. pomi* increases with age, the sensitivity of the young stages is greater than that of the older ones. At high concentration (LC95), values increase with stage; while for L4 and adults, lethality is variable (**Table 3** and **Figure 4**).

With regard to lethal concentrations, the hierarchical classification of the stages of *A. pomi* according to their sensitivity to crude OMW allows them to be classified in decreasing order L1 ≤ L2 ≤ L4 < Adults <L3. The nymphs L1, L2 and L4 were therefore more vulnerable to crude OMW than adults and L3 (**Figure 5**). In terms of efficiency, the application of 135 mg of crude OMW/L of water allows to reduce the level of aphid populations below the threshold adopted by producers (15–20% of infested shoots).

#### **4. Discussion**

#### **4.1 Physicochemical characteristics of OMW**

Olive oil extraction by traditional and three-phase processes generates huge amounts of vegetable waters rich in various organic and inorganic compounds during a short period of the year (October–March). OMW have a high biological and chemical oxygen demand values, as well as high contents of organic matter, suspended matter, inhibitor substances (phenolic compounds) and minerals,

*Valorization of Olive Mill Wastewater in the Control of* Aphis pomi *De Geer 1773… DOI: http://dx.doi.org/10.5772/intechopen.100016*

especially potassium, phosphorus, magnesium and calcium [58]. The values of the physicochemical parameters presented in this work are similar to those reported in Morocco (eg, [26, 59–62]). However, the chemical composition of OMW depends on the olive variety with the stage of maturity of the olives, the harvest period and the extraction techniques [63, 64]. Their physicochemical characteristics confer them the status of polluting substances posing serious environmental problems. Indeed, certain elements present in OMW are responsible for their harmful effects. Thus, the high concentration of phenolic compounds in vegetable waters generates phytotoxic effects [63]. Moreover, according to [65], heavy metals (Cu, Ni, Pb, Cr, Zn) affect the performance of anaerobic digestion by inhibiting microorganisms; indeed, during the production of biogas by the anaerobic digestion process from of the olive mill waste, the methanogenic bacteria are inhibited. The recovery, recycling, and reuse of these by-products are considered the best options for a sustainable water management program. Thus, their use as bio-pesticides is one of the valorization modalities that we have tried to apply in this work.

#### **4.2 Valorization of OMW in aphid control**

The damage caused by *A. pomi*, especially in nurseries and young plantations, force producers to treat frequently the pest with synthetic insecticides. The misuse of synthetic pesticides, however, raises safety and environmental issues [6–8]. At the same time, olive oil extraction by traditional and three-phase systems generates enormous volumes of vegetable waters, which are generally discharged into natural ecosystems [24–26]. To mitigate the undesirable effects of synthetic pesticides and at the same time those of OMW, it has proved to be imperative to replace synthetic aphicides by valorizing vegetable water in the management of *A. pomi*; this will contribute to crop protection while solving the environmental and health problems raised by the two categories of products. The treatment of the aphid with crude OMW has made it possible to prove their effectiveness. Thus, the crude vegetable waters tested against *A. pomi* were toxic to aphids. Indeed, the applied concentrations cause a variable mortality according to the stage and dependent on the concentration. The LC50 and LC95 vary from 27.17 to 45.59 and from 77.19 to 134.57 mg of crude OMW /L of water, respectively; the young stages were more vulnerable to vegetable waters than the older ones and adults. By high concentration, crude OMW equal the efficacy of the reference product, imidacloprid, used at the recommended dose in the field. In terms of efficiency, to reduce the level of populations below the threshold adopted by producers, ie, 15–20% of infested shoots, an average dose of 135 mg of crude OMW/L of water can satisfy the farmer to control aphids.

Various studies have evaluated the effectiveness of the OMW or their polyphenols in the management of insect pests on different rops. Thus, for example, the treatment of *Euphyllura olivina* Costa (Hemiptera, Psyllidae), olive psyllid, with polyphenols from OMW at a rate of 2 g of hydroxytyrosol/L of water in an olive grove, allowed to control 41.1% of larvae and 72% of adults. In contrast, with regard to eggs, the products was ineffective [44]. Against *A. pomi*, crude OMW or their polyphenols cause significantly higher mortalities than the controls used; their toxicity depends both on the stages of the insect and on the concentration of the products tested; the average LC50s are around 25.10 ml of crude OMW and 42.8 mg of polyphenols/L of distilled water. The toxicity caused by crude OMW depends in 67% to approximately 100% of cases on that of the polyphenols contained in them [45]. Spraying the larvae of *Potosia opaca* (Coleoptera, Scarabeidae), a date palm pest in Morocco, with crude OMW, allowed to obtain the same efficacy as with the reference insecticides (chlorpyriphos-ethyl or chlorpyriphos-ethy + cypermethrin) 19 days after treatment; vegetable waters are both toxic and affect the weight

of survivors; their toxicity depends directly on their polyphenol content [46]. Tested on the Mediterranean fruit fly (*Ceratitis capitata* (Wiedemann) (Diptera, Tephriditae), the polyphenolic fractions of OMW inhibit egg hatching and female fecundity without affecting larval development [49]. Overall, from all the studies cited in this paragraph, it emerges that the toxicity caused by OMW depends mainly on their polyphenol content (*op. Cit.*). In addition, although, the biochemical modes of action of OMW have not yet been elucidated in insects, the high levels of phenols present in vegetable water could block the transmission of nerve impulses [66, 67]. However, in this case, it is not excluded that the vegetable waters contained insecticides, in this case organophosphates and/or carbamates, used against the olive fly and which inhibit acetylcholinesterase (eg, [68]).
